The ability to 'exogenously' control the expression of genes in mammalian cells has been a powerful tool of biomedical research. In particular, gene regulation technology has played a major role in efforts to understand the role of specific gene products in fundamental biological processes and in development and disease states. The technology also offers the opportunity to have a major impact in a number of other areas of research and could even form an important basis for new therapeutic paradigms. However, due to limitations of existing systems, there is a need for the development of new methods for achieving gene regulation in order to realize the full potential of the technology. The focus of this grant proposal is the development of a new technology platform for gene regulation which involves the modulation of RNA self-cleavage, rather than transcription, as a means of controlling gene expression. The basis for the proposed research program are recent studies in our laboratory which demonstrate that incorporation of sequences encoding self-cleaving RNA motifs, ribozymes (rz), into a mammalian transcription unit can lead to the elimination of gene expression from the resulting transcript, due to efficient self-cleavage of the mRNA. In turn, we have shown that gene expression from such a configuration can be effectively 'induced' via inhibition of RNA self-cleavage using small molecule inhibitors. We believe that such a system for gene regulation may offer distinct advantages over existing systems for controlling gene expression and could have broad experimental and therapeutic application. The overall objective of the research program described here is to further understand the experimental parameters which govern efficient RNA self-cleavage in mammalian cells, and to apply that knowledge, in conjunction with development of new methodologies for the manipulation of rz self-cleavage and standard gene transfer technology, to the development of a widely applicable and 'user-friendly' technology platform for gene regulation. An important goal of the research program will be to establish a general methodology for the development of 'custom-designed' rz-based gene regulation systems tailored to respond to any specific small molecule or other molecular entity.